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Bioinformatics is a valuable tool to understand gene regulatory networks. Cis-regulatory elements (CREs) previously found in promoter regions are known to recruit transcription in signaling pathways. In this work it has been hypothesized to consider CREs as a family of related words that interact/bind to a family of related transcription factors, and thus have similar but distinct regulation patterns. A 1460 microarray gene expression collection was obtained via online databases to create a transcriptomic meta-dataset. A novel bioinformatic algorithm was applied to annotate all 65536 (64k) potential 8-letter CREs in the 500 bp upstream promoter region of all A. thaliana genes across the transcriptomic meta-dataset. Of the possible words, only 2,498 were significantly associated with a pattern of regulation in any of the 1,460 microarrays tested whereas the remaining motifs appeared not to be regulatory. Unique CREs were categorized into 4 regulatory types: inducer, suppressor, biregulator and insulator. A predicted protein protein interactome was created for an economically important plant Coffea canephora. Here, it has been hypothesized that evolutionary conservation of many core biological processes enable generation of predicted protein interactome for species with few resources other than sequenced genome. Of over 12,000 genes identified, 939 were predicted to have 4,587 interactions. Gene Ontology analysis revealed enrichment of processes conserved in all eukaryotes but depletion in unique plant processes. A third study was conducted to determine if homology modeling, evolutionary analysis, and structural evolution could determine key factors involved in function, and interaction specificity in Pus10 (EC 5.4.99.25) found in Archaea and Eukaryotes. Redundancy of Pus10 and the bacterial TrmA and TruB orthologs appear to have resulted in significant molecular evolution of Pus10 function. Neofunctionalization was identified in animal kingdom where thiouridine synthase, methylases and PSUSs (THUMP)-domain modification in early animal evolution coincides with appearance of TNF-related apoptosis-inducing ligand (TRAIL) apoptosis components. Subfunctionalization was identified for Thermococcales lineage of Archaea where a shorter forefinger-loop coincides with the loss of Ψ54 specificity as experimentally verified in P. furiosus. Absence of Pus10 was observed in Sulfolobus and higher fungi whereas in plant kingdom Pus10 function remains unknown with possible pseudogene in some lineages

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Magister Scientiae - MSc / Ischemic heart disease is a disease which is characterized by the reduced blood supply to the heart. According to WHO 2013, ischemic heart disease is one of the major causes of death globally. For this reason, it is imperative to search for methods whereby heart cells can be protected from cell death. The upregulation of heat shock proteins (Hsps) is one of the major techniques which can be used to protect the heart cells from Hsps cell death and improve the tolerance to ischemic stresses in various models. The increased expression of Hsps during heat shock pre-conditioning is regulated by heat shock transcription factors (HSFs). HSFs orchestrate the initiation of gene expression by binding to sequence motifs, known as cis-acting regulatory elements (CAREs). Since gene expression is regulated at a transcriptional level, it is expected that functionally related genes (e.g. heat shock response genes) might also be regulated by the same transcription factors (TFs). In this study an in silico approach was performed to identify the promoter sequences of 50 known heat shock responsive genes using Genomatix Software. This software was also used to identify transcription factor binding sites that are statistically over represented in the promoter sequences of these genes. The use of the Electrophoretic Mobility Shift Assay was included to confirm that protein cell lysates of stressed cells contain proteins (TFs) that bind to this sequence (SP1F_KLFS_01). Luciferase promoter reporter assay were also used to iii investigate the transcriptional activity of mutant promoter constructs in which the SP1F_KLFS_01 was mutated. SP1F_KLFS_01 is a ±25 base pair sequence that was identified in the promoter sequences of 19 heat shock responsive genes, including the well-known Hsp70 and Hsp90. This sequence is a potential binding site for two TFs, Specificity Protein-1 and Krueppel like TFs. Consequently, the aim of this study is to identify CAREs that are statistically over-represented in the promoter regions of heat shock response genes. In conclusion, in vitro experiments of this study did not support the findings of the in silico experiments, therefore additional methods should be implemented to expand the investigation for the involvement of cis-acting regulatory elements in the regulation of heat shock proteins in cardiomyocytes, prior to heat shock.

abstract: All biological processes like cell growth, cell differentiation, development, and aging requires a series of steps which are characterized by gene regulation. Studies have shown that gene regulation is the key to various traits and diseases. Various factors affect the gene regulation which includes genetic signals, epigenetic tracks, genetic variants, etc. Deciphering and cataloging these functional genetic elements in the non-coding regions of the genome is one of the biggest challenges in precision medicine and genetic research. This thesis presents two different approaches to identifying these elements: TreeMap and DeepCORE. The first approach involves identifying putative causal genetic variants in cis-eQTL accounting for multisite effects and genetic linkage at a locus. TreeMap performs an organized search for individual and multiple causal variants using a tree guided nested machine learning method. DeepCORE on the other hand explores novel deep learning techniques that models the relationship between genetic, epigenetic and transcriptional patterns across tissues and cell lines and identifies co-operative regulatory elements that affect gene regulation. These two methods are believed to be the link for genotype-phenotype association and a necessary step to explaining various complex diseases and missing heritability. / Dissertation/Thesis / Doctoral Dissertation Biomedical Informatics 2020

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京都大学 / 新制・課程博士 / 博士(医学) / 甲第24996号 / 医博第5030号 / 新制||医||1069(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 齊藤 博英, 教授 滝田 順子, 教授 小川 誠司 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

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